LOCUS       sequence-245483-        6119 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of SZ1-RGG-GFP-RGG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6119)
  AUTHORS   Schuster BS, Reed EH, Parthasarathy R, Jahnke CN, Caldwell RM, 
            Bermudez JG, Ramage H, Good MC, Hammer DA
  TITLE     Controllable protein phase separation and modular recruitment to 
            form responsive membraneless organelles.
  JOURNAL   Nat Commun. 2018 Jul 30;9(1):2985. doi: 10.1038/s41467-018-05403-1.
  PUBMED    30061688
REFERENCE   2  (bases 1 to 6119)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2018-07-30"; volume: "9(1):2985. doi"; pages: " 
            10.1038/s41467-018-05403-1"
COMMENT     Sequence Label: pET_SZ1-RGG-GFP-RGG
FEATURES             Location/Qualifiers
     source          1..6119
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     74..93
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     177..196
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        177..195
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    196..220
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             235..257
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             922..939
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             join(940..942,943..945,946..1656)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(940..942,943..945,946..1656)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(985..1006)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1246..1265)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1593..1614
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1663..1680
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             2191..2208
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2261..2279)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2275..2322
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2359..2814
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2446..2465)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2656..2677
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2929..3744)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2929..3744)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3652..3671
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3736..3755
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3897..4485
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4386..4405
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4639..4656
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(4827..5909)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5871..5890
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5910..5987)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 cggggcctgc caccataccc acgccgaaac aagcgctcat gagcccgaag tggcgagccc
       61 gatcttcccc atcggtgatg tcggcgatat aggcgccagc aaccgcacct gtggcgccgg
      121 tgatgccggc cacgatgcgt ccggcgtaga ggatcgagat ctcgatcccg cgaaattaat
      181 acgactcact ataggggaat tgtgagcgga taacaattcc cctctagaaa taattttgtt
      241 taactttaag aaggagatat acatatgaac ttggtagccc agcttgagaa cgaagttgcg
      301 agtctggaaa acgaaaacga gacgttgaag aaaaagaact tgcacaagaa agatttgatt
      361 gcctacttgg aaaaggaaat tgccaatctg cgtaaaaaga ttgaggaagg tatggaatcc
      421 aatcaatcca ataacggtgg ctcgggtaat gccgctctga atcgtggtgg ccgctacgtg
      481 ccgccgcatc tgcgtggtgg cgatggtggt gcagcagcag ctgcatctgc tggcggtgat
      541 gaccgtcgcg gcggtgcggg cggtggcggt tatcgtcgcg gcggtggcaa tagtggtggc
      601 ggtggcggtg gcggttatga tcgtggttac aatgacaacc gtgatgaccg cgataaccgt
      661 ggcggttccg gcggttacgg tcgtgatcgt aattatgaag accgtggtta caacggcggt
      721 ggcggtggcg gtggcaatcg tggctataac aataaccgtg gtggcggtgg cggtggctac
      781 aatcgccagg atcgtggtga cggtggcagc tctaactttt cgcgcggtgg ctataataac
      841 cgtgatgaag gcagcgacaa ccgcggtagt ggccgttcct acaataacga tcgtcgcgac
      901 aatggtggcg atggcgaatt cttagtacct cgcgggtcaa tggtgagcaa gggcgaggag
      961 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
     1021 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc
     1081 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac
     1141 ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
     1201 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
     1261 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     1321 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac
     1381 agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag
     1441 atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc
     1501 cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccgcc
     1561 ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc
     1621 gccgggatca ctctcggcat ggacgagctg tacaaggaat tcttagtacc tcgcgggtca
     1681 atggaatcca atcaatccaa taacggtggc tcgggtaatg ccgctctgaa tcgtggtggc
     1741 cgctacgtgc cgccgcatct gcgtggtggc gatggtggtg cagcagcagc tgcatctgct
     1801 ggcggtgatg accgtcgcgg cggtgcgggc ggtggcggtt atcgtcgcgg cggtggcaat
     1861 agtggtggcg gtggcggtgg cggttatgat cgtggttaca atgacaaccg tgatgaccgc
     1921 gataaccgtg gcggttccgg cggttacggt cgtgatcgta attatgaaga ccgtggttac
     1981 aacggcggtg gcggtggcgg tggcaatcgt ggctataaca ataaccgtgg tggcggtggc
     2041 ggtggctaca atcgccagga tcgtggtgac ggtggcagct ctaacttttc gcgcggtggc
     2101 tataataacc gtgatgaagg cagcgacaac cgcggtagtg gccgttccta caataacgat
     2161 cgtcgcgaca atggtggcga tggcctcgag caccaccacc accaccactg agatccggct
     2221 gctaacaaag cccgaaagga agctgagttg gctgctgcca ccgctgagca ataactagca
     2281 taaccccttg gggcctctaa acgggtcttg aggggttttt tgctgaaagg aggaactata
     2341 tccggattgg cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     2401 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     2461 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     2521 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     2581 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     2641 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     2701 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     2761 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttaggtgg
     2821 cacttttcgg ggtacccagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     2881 ttatttttct aaatacattc aaatatgtat ccgctcatga attaattctt agaaaaactc
     2941 atcgagcatc aaatgaaact gcaatttatt catatcagga ttatcaatac catatttttg
     3001 aaaaagccgt ttctgtaatg aaggagaaaa ctcaccgagg cagttccata ggatggcaag
     3061 atcctggtat cggtctgcga ttccgactcg tccaacatca atacaaccta ttaatttccc
     3121 ctcgtcaaaa ataaggttat caagtgagaa atcaccatga gtgacgactg aatccggtga
     3181 gaatggcaaa agtttatgca tttctttcca gacttgttca acaggccagc cattacgctc
     3241 gtcatcaaaa tcactcgcat caaccaaacc gttattcatt cgtgattgcg cctgagcgag
     3301 acgaaatacg cgatcgctgt taaaaggaca attacaaaca ggaatcgaat gcaaccggcg
     3361 caggaacact gccagcgcat caacaatatt ttcacctgaa tcaggatatt cttctaatac
     3421 ctggaatgct gttttcccgg ggatcgcagt ggtgagtaac catgcatcat caggagtacg
     3481 gataaaatgc ttgatggtcg gaagaggcat aaattccgtc agccagttta gtctgaccat
     3541 ctcatctgta acatcattgg caacgctacc tttgccatgt ttcagaaaca actctggcgc
     3601 atcgggcttc ccatacaatc gatagattgt cgcacctgat tgcccgacat tatcgcgagc
     3661 ccatttatac ccatataaat cagcatccat gttggaattt aatcgcggcc tagagcaaga
     3721 cgtttcccgt tgaatatggc tcataacacc ccttgtatta ctgtttatgt aagcagacag
     3781 ttttattgtt catgaccaaa atcccttaac gtgacctagg tcatgaccaa aatcccttaa
     3841 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     3901 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     3961 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     4021 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     4081 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     4141 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     4201 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     4261 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     4321 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     4381 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     4441 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     4501 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     4561 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     4621 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg ccataatcca
     4681 tgataaagaa gacagtcata agtgcggcga cgatagtcat gccccgcgcc caccggaagg
     4741 agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc taatgagtga
     4801 gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     4861 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgcc
     4921 agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt caccgcctgg
     4981 ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt
     5041 ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc gtatcccact
     5101 accgagatat ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc
     5161 gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt cagcatttgc
     5221 atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc tatcggctga
     5281 atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc cgagacagaa
     5341 cttaatgggc ccgctaacag cgcgatttgc tggtgaccca atgcgaccag atgctccacg
     5401 cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt ctggtcagag
     5461 acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat ggcatcctgg
     5521 tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc
     5581 gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac gctggcaccc
     5641 agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg cagggccaga
     5701 ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg
     5761 ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa
     5821 acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc ggcatactct
     5881 gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact ctcttccggg
     5941 cgctatcatg ccataccgcg aaaggttttg cgccattcga tggtgtccgg gatctcgacg
     6001 ctctccctta tgcgactcct gcattaggaa gcagcccagt agtaggttga ggccgttgag
     6061 caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg cccaacagtc ccccggcca
//
